The present invention relates to methods for reducing free radical levels in mammals. In a particular aspect, the present invention relates to methods for treating free radical overproduction in mammals by administration of physiologically active dithiocarbamate compounds which non-covalently bind free radicals in hosts afflicted with a variety of disorders. In a further aspect, the present invention relates to methods for treating free radical overproduction in mammals by administration of physiologically compatible dithiocarbamate-containing free radical scavengers in hosts afflicted with a variety of disorders.
Free-radicals are chemical species having one or more unpaired electrons. Such species are formed by a variety of processes including photolysis, thermal homolysis, and radical-forming redox reactions. Nitric oxide, for example, is formed by the enzymatic oxidation of:L-arginine to citrulline via the action of nitric oxide synthase. Radical formation can also proceed via a variety of chain reactions. For example, the reduction of molecular oxygen by one electron gives rise to superoxide radical anion. Addition of a second electron to superoxide produces peroxide ion, which lacks an unpaired electron and is thus not a radical. Peroxide ion, however, will almost immediately protonate hydrogen peroxide at physiological pH, yielding a molecule of water and a molecule of hydroxyl radical, the strongest known oxidant produced in biological systems.
Free radicals are produced in conjunction with a variety of normal biological processes, such as mitochondrial electron transfer, and can play important roles in normal physiology. Because of their high reactivity, overproduction or inappropriate production of free radical species can be detrimental. For example, generation of superoxide radical is associated with reperfusion injury following stroke or acute myocardial infarction (McCord, N. Engl. J. Med. 312:159-163, 1985), and hydroxyl radicals are associated with a variety of neurodegenerative disorders (Evans, Br. Med. Bull. 49:577-587, 1993).
The majority of treatments for excess free radical production available to date involve the use of enzymatic inhibitors to reduce radical production, or the use of protein-based antioxidants, such as superoxide dismutase, to neutralize and reduce the potential cytotoxic effects of free radicals. Both of these approaches have proved unsatisfactory, in particular because by the time a patient presents with symptoms and can be treated, substantial amounts of free radicals have already been produced.
The use of dithiocarbamates in medical applications has been proposed in. limited circumstances. For example, dithiocarbamates have been used clinically for treatment of heavy metal intoxication. See, for example, F. W. Sunderman in Ann Clin Lab Sci 9(1):1-10 (1979); and M. M. Jones et al., in Fundamental Appl. Toxicology 19:432-437 (1992). Dithiocarbamates have also been used to alleviate renal toxicity associated with cis-platinum chemotherapy. See, for example, R. Qazi et al., in J Natl Cancer Inst 80:1486-1488 (1988); and D. R. Gandara et al., in J Clin Oncol 13:490-496 (1995). A specific dithiocarbamate, diethyldithiocarbamate, has been shown to inhibit HIV progression. This has led to clinical trials for the use of this compound in the treatment of AIDS patient populations (see, for example, E. Reisinger et al., in Lancet 335:679-682 (1990)).
Diethyldithiocarbamate has also been the subject of a limited study on potential in vitro antioxidants. See Ames, et. al. Free Rad. Res., 24(6), p. 461, (1996). This study addresses only the in vitro reaction chemistry of diethyldithiocarbamate with HOCI and hydroxyl radical, but is conspicuously silent on the use of this or any other dithiocarbamate for the in vivo reduction of free radical levels.
Dithiocarbamates such as pyrrolidine dithiocarbamate have been determined to be potent inhibitors of nuclear factor kappa B (NFKB) in intact cells (see, for example, R. Schreck et al., in J Exp Med 175:1181-1194 (1992). In addition, NFKB has also been shown to up-regulate the expression of cell adhesive molecules, including the vascular cell adhesive molecule-1 (VCAM-1; see, for, example, M. F. Iademarco et al., in J Biol Chem 267:16323-16329 (1992)). Interestingly, in view of these known effects of dithiocarbamates on NFKB, Medford et al. propose the use of dithiocarbamates to treat cardiovascular diseases mediated by VCAM-1, through the inhibition of the NFKB pathway (see U.S. Pat. No. 5,380,747).
Accordingly, there is still a need in the art for processes to reduce free radical levels in vivo. In addition, there is also a need in the art for reagents useful for treatment of the cytotoxic effects associated with free radical overproduction, such as occurs in reperfusion injury, neurodegenerative disorders, and a variety of other diseases and conditions. The present invention addresses these and related needs.
In accordance with the present invention, methods have been developed for the in vivo reduction of free radical levels in subjects in need thereof. In contrast to treatment approaches described in the prior art, the present invention employs a scavenging approach whereby overproduced free radical is bound in vivo to a suitable free radical scavenger. The resulting complex renders the free radical harmless, and is eventually excreted in the urine of the host. Further in accordance with the present invention, there have been developed compositions and formulations useful for carrying out the above-described methods.
An exemplary free radical scavenger contemplated for use in the practice of the present invention is a dithiocarbamate-ferrous iron complex. This complex binds non-covalently to free radicals, forming a stable, water-soluble dithiocarbamate-iron-free radical complex.
The present invention relates to methods for reducing in vivo levels of free radicals as a means of treating subjects afflicted with inflammatory and/or infectious disease, reperfusion injury, neurodegerative disorders, and the like. Suitable free radical scavengers are administered to a host in need of such treatment; these scavengers interact with in vivo produced free radical, forming a stable non-covalently bound free radical-containing complex. Whereas un-complexed free radicals are highly reactive chemical species, non-covalently bound free radical-containing complexes are not. The free radical-containing complex is then filtered through the kidneys, concentrated in the urine, and eventually excreted by the subject, thereby reducing in vivo free radical levels.